The overall objectives of our research during the previous funding period was to fain further insight about the genetic regulation of the CD11/CD18 family of beta2 integrins and to determine the functional relevance of these adhesion-promoting glycoproteins in the cellular acute inflammatory response. As an outgrowth of these efforts, we have accumulated data that suggest that the Mo1 (CD11b/CD18) integrin glycoprotein not only serves as a traditional receptor for well-defined ligands such as ICAM-1 (CD54) and C3bi, but also serves as a mechanotransducer of signals triggered by the ligation of other phagocyte receptors such as FcgammaRIIIB (CD16b) and the urokinase receptor, uPA-R (CD87) (GPI-anchored molecules that lack cytoplasmic domains). Specifically, we have demonstrated a physical and functional linkage between Mo1 and FcgammaRIIIB and between Mo1 and uPA-R, and it is the goal of this application to more fully characterize the regulation, structural requirements, and functional consequences of these interactions. The specific aims of the proposed research plan are as follows: (1) Extend our characterization of the physical and functional association between Mo1 and FcgammaRIIIB by: (a) assessing the conditions (resting or activated) under which physical linkages between Mo1 and FcgammaRIIIB occur on the phagocyte membrane (quantitating the extent to which linkages exist by resonance energy transfer [RET]), immunogold electron microscopy, and co-precipitation after bifunctional chemical cross-linking, (b) determining the contribution of specific FcgammaRIIIB oligosaccharides to the formation and maintenance of physical and functional associations with Mo1, (c) evaluating the role of Mo1 as a transducer of FcgammaRIIIB-triggered signals leading to cellular effector responses (in transfectant cell lines expressing mutant forms of the Mo1 glycoprotein), and (d) determining the extent to which a physical linkage with Mo1 influences the affinity of FcgammaRIIIB for Ig. (2) Substantiate the functional significance of a similar linkage between Mo1 and uPA-R, and apply the same strategies as outlined in Aim 1 to define the regulation, structural requirements, and functional consequences of this interaction. (3) Explore the possible existence of multiple receptor- receptor interactions involving Mo1, FcgammaRIIIB, uPA-R, FcgammaRII, and the receptor for formyl peptides (FPR) on the neutrophil membrane by (a) RET studies to define the conditions under which receptor interactions may occur (based upon molecular proximity), and (b) transfection studies to determine the physiologic significance of possible linkages among these five receptor structures. It is the goal of this investigation to test the hypothesis that cooperative interactions among neutrophil receptors govern the neutrophil response to certain external stimuli and contribute to host immunity.